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Objective. This study is designed to find out the molecular targets of effective Chinese medicine Ziyin Mingmu pills (ZMPs) in treating age-related macular degeneration (AMD) based on network pharmacology and experimental data. Methods. A comprehensive network pharmacology strategy that consists of three sequential modules (drug-disease target molecular docking, enrichment analysis, and external verification) was carried out to identify potential targets of ZMPs acting on AMD. Results. The active ingredients of ZMPs targeting 66 genes have effects on the process of AMD. GO and KEGG pathway enrichment analyses suggested that response to oxidative stress, regulation of angiogenesis, and lipid and atherosclerosis might serve as the most important signaling pathways in ZMPs for AMD treatment. Combined with the GSE29801 dataset for further analysis, two key genes, EGFR and VEGFA, were identified. Immune infiltration analysis showed that there was a strong association between EGFR and immune cell content. In addition, images were acquired following 24 h in the scratch experiment showed that ZMPs can reduce the percentage of wound healing distance. The Western blot assay found that ZMPs increased the expression of EGFR and decreased the expression of VEGFA. Conclusion. This study sheds light on some mechanisms of ZMP therapy for AMD, particularly the effect of ZMP on the oxidative stress in RPE and cell survival and angiogenesis in AMD. We propound ZMPs as a promising strategy to intervene in the process of AMD. [ABSTRACT FROM AUTHOR]

Multidrug resistance (MDR) is a critical reason for cancer chemotherapy failure. Babaodan (BBD) is a famous traditional Chinese patent medicine reported to have antigastric cancer activity. However, the roles and molecular mechanisms of the reversal of MDR of gastric cancer by BBD have not been well described until now. Therefore, the purpose of this study was to elucidate further the role of BBD in reversing the MDR of gastric cancer cells and its specific regulatory mechanism via in vitro experiments. To verify our results, MTT, Doxorubicin (DOX) staining, Rhodamin123 (Rho123) staining, DAPI staining, Annexin V-FITC, propidium iodide (PI), Cyto-ID, and western blot assays were performed. To determine whether BBD triggers apoptosis and autophagy through the PI3K/AKT/mTOR signaling, we also applied 3-methyladenine (3-MA), chloroquine (CQ), and 740Y-P (an activator of PI3K). The results showed that BBD reversed the MDR and induced apoptosis and autophagy of SGC7901/DDP cells. Pathway analyses suggested BBD inhibits PI3K/AKT/mTOR pathway activity and subsequent apoptosis-autophagy induction. Inhibition of autophagy with 3-MA and chloroquine (CQ) was performed to confirm that BBD promoted autophagy. PI3K agonist, 740Y-P, further verified BBD inhibition of PI3K/AKT/mTOR pathway activation. In conclusion, BBD may reverse the MDR of gastric cancer cells, induce apoptosis, and promote autophagy via inactivation of the PI3K/AKT/mTOR signaling pathway. [ABSTRACT FROM AUTHOR]

The use of 5-fluorouracil (5-FU) has been proven benefits, but it also has adverse events in colorectal cancer (CRC) chemotherapy. In this study, we explored the mechanism of 5-FU resistance by bioinformatics analysis of the NCBI public dataset series GSE81005. Fifteen hub genes were screened out of 582 different expressed genes. Modules of the hub genes in protein-protein interaction networks gathered to TOP2α showed a decrease in HCT-8 cells but an increase in 5-FU-resistant HCT-8/5-FU cells with 5-FU exposure. Downregulation of TOP2α with siRNA or miR-494 transfection resulted in an increase of cytotoxicity and decrease of cell colonies to 5-FU for HCT-8/5-FU cells. Moreover, we found that an ethanol extract of Spica Prunellae (EESP), which is a traditional Chinese medicine with clinically beneficial effects in various cancers, was able to enhance the sensitivity of 5-FU in HCT-8/5-FU cells and partly reverse the 5-FU resistance effect. It significantly helped suppress cell growth and induced cell apoptosis in HCT-8/5-FU cells with the expression of TOP2α being significantly suppressed, which increased by 5-FU. Consistently, miR-494, which reportedly regulates TOP2α, exhibited reverse trends in EESP/5-FU combination treatment. These results suggested that Spica Prunellae may be beneficial in the treatment of 5-FU-resistant CRC patients. [ABSTRACT FROM AUTHOR]

Objective: To evaluate the effect of Pien Tze Huang (片仔癀, PZH) on breast cancer chemoresistance and related epithelial-mesenchymal transition (EMT) and investigate the underlying mechanisms. Methods: 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) assay was used to determine the cell viability. Adriamycin (ADR) staining observed by fluorescence microscope was performed to detect the accumulation of ADR. Transwell assay was used to analyze the cell migration and invasion. Western-blot was performed to detect the protein expression of related genes. Results: MCF-7/ADR cells were resistant to ADR treatment, and PZH treatment inhibited the viability of MCF-7/ADR cells in a dose-dependent manner. PZH treatment also increased the intercellular accumulation of ADR and down-regulated the expression of ABCG2 and ABCB1 in MCF-7/ADR cells (P<0.05). In addition, PZH treatment inhibited EMT, migration and invasion of MCF-7/ADR cells (P<0.05). Moreover, PZH suppressed activation of transforming growth factor β1 (TGF-β) signaling in MCF-7/ADR cells (P<0.05). Conclusion: PZH treatment can effectively overcome chemoresistance via down regulating ABCG2, ABCB1 and inhibit EMT in ADR resistant human breast cancer cells via suppression of the TGF-β1 pathway. [ABSTRACT FROM AUTHOR]

Summary: Acute myeloid leukemia (AML) is the most common form of acute leukemia with dismal long-term prognosis with age. The most aggressive subtype of AML is MLL-AML that is characterized by translocations of the mixed-lineage leukemia gene (MLL) and resistance to conventional chemotherapy. Cyclin dependent kinase 9 (CDK9) plays a crucial role in the MLL-driven oncogenic transcription, and hence, inhibiting activity of CDK9 has been proposed as a promising strategy for treatment of AML. We investigated the therapeutic potential of CDKI-73, one of the most potent CDK9 inhibitors, against a panel of AML cell lines and samples derived from 97 patients. CDKI-73 induced cancer cells undergoing apoptosis through transcriptional downregulation of anti-apoptotic proteins Bcl-2, Mcl-1 and XIAP by majorly targeting CDK9. Contrastively, it was relatively low toxic to the bone marrow cells of healthy donors. In MV4–11 xenograft mouse models, oral administration of CDKI-73 resulted in a marked inhibition of tumor growth (p < 0.0001) and prolongation of animal life span (P < 0.001) without causing body weight loss and other overt toxicities. The study suggests that CDKI-73 can be developed as a highly efficacious and orally deliverable therapeutic agent for treatment of AML. [ABSTRACT FROM AUTHOR]

Objective: To investigate the effects of ethanol extract of Patrinia scabiosaefolia (EEPS) on chemo-resistance of colorectal cancer cells (CRC) and explore the possible molecular mechanisms.Methods: 5-fluorouracil (5-FU)-resistant human colorectal carcinoma cell line (HCT-8/5-FU) and its parental cells HCT-8 were treated with EEPS (0, 0.25, 0.50, 1 or 2 mg/mL), or 5-FU (0, 100, 200, 400, 800 or 1600 μmol/L). The 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay was performed to evaluate the cell viability. Cell density was observed by phase-contrast microscope, cell counting and colony formation assay were used to determine the cell proliferation of HCT-8/5-FU cells treated with 0, 0.5, 1 or 2 mg/mL EEPS. Cell apoptosis was determined by Hoechst staining. Western-blot was performed to detect the phosphorylation of AKT as well as the protein expression level of B-cell CLL/lymphoma 2 (Bcl-2) and Bcl-2-associated X protein (Bax).Results: Compared with HCT-8 cells, MTT assay results indicated that HCT-8/5-FU cells were resistant to 5-FU treatment (P<0.05), and sensitive to EEPS treatment (P>0.05). Moreover, compared with untreated HCT-8/5-FU cells, 1 and 2 mg/mL of EEPS treatment significantly reduced cell density, cell number, inhibited cell survival (P<0.05), and induced apoptosis in HCT-8/5-FU cells. Furthermore, 1 and 2 mg/mL of EEPS significantly decreased the phosphorylation level of p-AKT and Bcl-2 protein expression, and increased the expression of Bax protein (P<0.05).Conclusion: EEPS is a promising therapeutic agent that may overcome chemo-resistance in cancer cells, likely through suppression of the AKT pathway and promotion of cancer cell apoptosis. [ABSTRACT FROM AUTHOR]

Hypoxia-induced angiogenesis plays an important role in the development and metastasis of solid tumors and is highly regulated by HIF-1α/VEGF-A pathway. Therefore, inhibiting tumor angiogenesis via suppression of HIF-1α/VEGF-A signaling represents a promising strategy for anticancer treatment. As a traditional Chinese medicine formula, Pien Tze Huang (PZH) has long been used as a folk remedy for cancer in China and Southeast Asia. Previously, we reported that PZH inhibits colorectal cancer (CRC) growth both in vivo and in vitro. To elucidate the antitumor mechanisms of PZH, in the present study we used human umbilical vein endothelial cells (HUVEC) and colorectal carcinoma HCT-8 cells to evaluate the effects of PZH on hypoxia-induced angiogenesis and investigated the underlying molecular mechanisms. We found that PZH could inhibit hypoxia-induced migration and tube formation of HUVEC cells in a dose-dependent manner, although the low concentrations of PZH had no effect on HUVEC viability. Moreover, PZH inhibited hypoxia-induced activation of HIF-1α signaling and the expression of VEGF-A and/or VEGFR2 in both HCT-8 and HUVEC cells. Collectively, our findings suggest that PZH can inhibit hypoxia-induced tumor angiogenesis via suppression of HIF-1α/VEGF-A pathway. [ABSTRACT FROM AUTHOR]

Objective: To investigate the molecular mechanisms by which Qianliening Capsule (前列宁胶囊, QC) treats benign prostatic hyperplasia (BPH). Methods: Human prostate stromal cell line WPMY-1 was treated with 0, 1, 3 and 5 mg/mL of QC for 24, 48 and 72 h, respectively, in the presence of 10 ng/mL basic fibroblast growth factor (bFGF). The viability of WPMY-1 cells was determined by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Cell morphology was observed by phase-contrast microscopy. 4′,6-diamidino-2-phenylindole (DAPI) staining and fluorescence activated cell sorting (FACS) analysis with Annexin-V/propidium iodide (PI) staining were performed to determine cell apoptosis. The loss of mitochondrial membrane potential was examined by FACS analysis with 5,5′,6,6′-tetrachloro-1,1′,3,3′-tetraethylbenzimidazolyl-carbocyarine iodide (JC-1) staining. Activation of caspase-3 and -9 was evaluated by colorimetric assay. The mRNA and protein expression levels of Bcl-2 and Bax were measured by reverse transcription polymerase chain reaction (RT-PCR) and Western blotting, respectively. Results: Upon bFGF stimulation, the viability of WPMY-1 cells was increased to 122%-118% compared with the control cells ( P <0.05). However, treatment with 1-5 mg/mL of QC for 24, 48 and 72 h decreased the viability of bFGF-stimulated cells to 80%-92%, 59%-82%, 36%-62% compared with the untreated cells ( P <0.05). In addition, QC treatment reduced WPMY-1 cell density in a dose-dependent manner. Moreover, QC treatment dose-dependently induced the loss of plasma membrane asymmetry, the nuclear condensation and fragmentation, collapse of mitochondrial membrane potential, activation of caspase-9 and caspase-3, and increase of pro-apoptotic Bax/Bcl-2 ratio. Conclusion: Promoting mitochondrion-dependent apoptosis of prostate stromal cells might be one of the mechanisms by which QC treats BPH. [ABSTRACT FROM AUTHOR]

Backgrounds: LncRNA long intergenic non-protein coding RNA p53 induced transcript (LINC-PINT) is downregulated in multiple types of cancer cells. The authors explored the possible involvement of LINC-PINT in esophageal squamous cell carcinoma (ESCC). Materials and Methods: Sixty-two patients with early-stage ESCC were included in this study. Reverse transcription-polymerase chain reaction (RT-PCR) was used to detect gene expression in plasma from ESCC patients and healthy controls. Diagnostic analysis was performed by receiver operating characteristic (ROC) curve. Transfections were performed to analyze gene interactions. Cell invasion and migration were analyzed by Transwell assays. Results: Plasma LINC-PINT was downregulated and microRNA (miRNA)-21 was upregulated in early-stage ESCC patients. Diagnostic analysis by ROC curve revealed that downregulation of lncRNA LINC-PINT distinguished ESCC patients from healthy controls. Plasma levels of LINC-PINT and miRNA-21 were negatively correlated in ESCC patients. After surgical resection, only local recurrence was observed during 3-years of follow-up. LINC-PINT expression was further downregulated in recurrent patients but not in nonrecurrent patients. ROC curve analysis revealed that plasma levels of LINC-PINT at 12 months before recurrence can be used to distinguish ESCC patients from healthy controls. Overexpression of LINC-PINT could inhibit the expression of miRNA-21 in human ESCC cells, whereas LINC-PINT expression was not altered by miR-21 overexpression. Conclusion: Therefore, downregulation of LINC-PINT participated in the recurrence of ESCC possibly by interacting with miRNA-21. [ABSTRACT FROM AUTHOR]

A physiologically relevant glioma tumor model is important to the study of disease progression and screening drug candidates. However, current preclinical glioma models lack the brain microenvironment, and the established tumor cell lines do not represent glioma biology and cannot be used to evaluate the therapeutic effect. Here, we reported a real-time integrated system by generating 3D ex vivo cerebral organoids and in vivo xenograft tumors based on glioma patient-derived tissues and cells. Our system faithfully recapitulated the histological features, response to chemotherapy drugs, and clinical progression of their corresponding parental tumors. Additionally, our model successfully identified a case from a grade II astrocytoma patient with typical grade IV GBM features in both organoids and xenograft models, which mimicked the disease progression of this patient. Further genomic and transcriptomic characterization was associated with individual clinical features. We have demonstrated the "GBM-&Normal-like" signature to predict prognosis. In conclusion, we developed an integrated system of parallel models from patient-derived glioma cerebral organoids and xenografts for understanding the glioma biology and prediction of response to chemotherapy drugs, which might lead to a new strategy for personalized treatment for this deadly disease. [ABSTRACT FROM AUTHOR]

Traditional Chinese Medicine (TCM) is being increasingly used worldwide due to its diverse efficacy and relatively low side effects. Babao Dan (BBD) is a well-known TCM formula that is currently used for the effective treatment of various cancers, however its underlying molecular mechanism remains unknown. Tumor growth and tumor recurrence are characterized by two distinct populations of cells, namely the well-differentiated cancer cells composing the majority of tumor bulk, and cancer stem cells (CSCs) involved in tumor relapse, which are both strongly associated with excessive activation of Wnt/β-catenin signaling. Our study aims to elucidate the underlying molecular mechanisms associated with the anti-tumor proliferative effects of Babao Dan (BBD). We used a hepatoblastoma cell line HepG2 with stem cell-like traits that harbors a constitutively active mutant of β-catenin in order to study the anti-tumor ability of BBD via targeting Wnt/β-catenin signaling. BBD robustly attenuated both the intrinsic and extrinsic activation of Wnt/β-catenin pathway in HepG2 hepatoblastoma cells, as well as Wnt target genes. Moreover, BBD significantly inhibited both the proliferation of well-differentiated cancer cells, as well as the stem-like property of CSCs as evidenced by EpCAM, a Wnt target gene and a novel marker of cancer cell stemness. In addition, mice administered with BBD using HepG2 cell line derived xenograft model had marked reductions in tumor size and weight, as well as significantly decreased expressions of Wnt target genes and cancer cell stemness. Our findings elucidated the underlying molecular mechanisms associated with the robust anti-tumor effects of BBD via potent inhibition of Wnt/β-catenin signaling, and implicate its use in the clinical treatment of cancers. [Display omitted] • BBD inhibits cancer cell proliferation and stem-like property of cancer stem cells. • BBD attenuates both intrinsic and extrinsic activation of Wnt/β-catenin pathway. • BBD inhibits EpCAM expression, a Wnt target gene and marker of cancer cell stemness. • Robust anti-tumor effects of BBD was via potent inhibition of Wnt/β-catenin signaling. [ABSTRACT FROM AUTHOR]

MicroRNA (miR)-93 has been proven to mediate the initiation and progression of colorectal carcinoma (CRC); however, the mechanisms by which miR-93 mediates CRC development need deeper elucidation. The present study is designed to investigate the association between miR-93 and high mobility group box 3 (HMGB3), as well as the functions of miR-93, in CRC. miR-93 expression was quantified by RT-qPCR. CRC cells were transfected or cotransfected with miR-93 mimic, miR-93 inhibitor, pcDNA3.1-HMGB3 and sh-HMGB3, and then the proliferative, migratory and invasive capacities were detected in addition to the apoptotic rate. Western blotting assessed the expression levels of HMGB3, PI3K, p-PI3K, AKT and p-AKT. The interaction between miR-93 and HMGB3 was identified. In CRC tissues, miR-93 was downregulated and HMGB3 was upregulated. LOVO and SW480 cells transfected with miR-93 mimic exhibited reduced proliferation, invasion and migration as well as increased apoptosis. The ratios of p-PI3K/PI3K and p-AKT/AKT were declined after miR-93 mimic was introduced into the CRC cell lines. miR-93 negatively downregulated HMGB3, and introduction of pcDNA3,1-HMGB3 could counteract, in part, the inhibitory effects of miR-93 on the malignant properties of CRC cells as well as the ratios of p-PI3K/PI3K and p-AKT/AKT. miR-93 targeted HMGB3 to block the activation of the PI3K/AKT pathway and thus enhance CRC cell apoptosis. [ABSTRACT FROM AUTHOR]

• A porcine alveolar macrophage cell line stably expressing CD163 was established using lentivirus integration system. • The stable cell line expressing CD163 was susceptible to PRRSV infection and yielded high titer of progeny virus. • After virus infection the new cell line demonstrated cytokine secretion characteristics similar to primary alveolar macrophages. The in vitro investigation of cytokine secretion induced by porcine reproductive and respiratory syndrome virus (PRRSV) requires porcine alveolar macrophages (PAMs) and their interaction with immunocytes. However, immortalized monoclonal PAMs (mPAMs) are non-permissive for PRRSV infection. The porcine CD163 receptor isolated from primary PAMs (pPAMs) confers susceptibility to PRRSV infection; thus, this approach could be used to establish a novel cell line to facilitate the exploration of PRRSV infection kinetics. Here, we amplified the coding region of the CD163 gene from pPAMs and integrated it into an mPAM line using a lentivirus expression system. After verification, the monoclonal PAM cell line stably expressing CD163 (mPAM-CD163-GFP) was infected with either the highly pathogenic PRRSV strain JXA1 or the classical PRRSV strain SD1, which produced high infectious titers of progeny virus reaching > 109 copies/mL or a 50 % tissue culture infective dose of 105.5 over at least 100 cell generations. We also investigated cytokine and Toll-like receptor expression in infected mPAM-CD163-GFP cells and pPAMs. The mPAM-CD163-GFP cell line showed similar patterns of viral replication and cytokine secretion compared with pPAMs, so it may be extremely useful for replacing primary cells for in vitro investigations of the mechanisms of cytokine secretion and interactions between PRRSV-infected PAMs and immunocytes. [ABSTRACT FROM AUTHOR]